Co-immunoprecipitation of the small DUF1127 protein RSP_6037xFLAG and pRK0557_3xFLAG for RNA interaction

Many protein domains are conserved among different classes and are found in numerous species, but their function remains obscure. Data bases list more than 17,000 DUF1127 proteins but to date no biological function could be assigned to any of these proteins. This study demonstrates that the 71 amino acid DUF1127 protein CcaF1 from the alphaproteobacterium Rhodobacter sphaeroides participates in the maturation of the CcsR sRNAs that are processed from the 3´ UTR of the ccaF mRNA and have a role in the oxidative stress defense. CcaF1 also affects the abundance of several other sRNAs but also of tRNAs, rRNAs and several mRNAs. Co-immunoprecipitation revealed direct binding of CcaF1 to many cellular RNAs of different type and an effect of CcaF1 on the stability of selected RNAs was demonstrated. Thus, the widely distributed DUF1127 domain can mediate RNA-binding and affect stability of its binding partners and consequently the bacterial transcriptome. Overall design: Protein-RNA interaction was analyzed by co-immunoprecipitation. The strains Rhodobacter sphaeroides 2.4.1 pRK6037_3xFLAG (control pRK6037) and pRK0557_3xFLAG (control pRK0557) were used. RNA was isolated and sequenced.